Submission Procedure
Preparing your sample for submission
P r i m e r s
Designing your own primer:
- Primers should be between 18 and 30 base pairs
- Tm should be between 50°C and 70°C
- Avoid having runs of the same base
- Avoid primers with secondary structure or which can dimerize
Primers available from the facility:
D N A Te m p l a t e
The template may be a pcr product, plasmid, BAC or cosmid.
Q u a l i t y
The quality of the template is one of the primary determinants of the quality of sequence. Please take the time to purify your sample. Commercially available kits, such as Qiagen’s, yield great results. We do not recommend alkaline lysis as purifying method.
Q u a n t i t y
Please submit one template with one primer per tube. Should you wish to sequence the same template with multiple primers, please submit multiple reactions.
The template+primer+water should be submitted in a total volume of 12 µls.
|
Template |
Primer |
Water |
PCR Product
|
50 – 150 ng |
3.2 pmoles |
To 12 µls |
Plasmid (<9kb
|
600 - 800 ng |
3.2 pmoles |
To 12 µls
|
Plasmid (>9kb
|
1.0 µg |
25 pmoles |
To 12 µls |
BAC
|
1.0 – 1.5 µgs |
13 pmoles |
To 12 µls |
Dropping off your samples
Please drop off your samples in the DNA sequencing freezer located in SL-429.
If you do not have access to the Slosberg building, please use phone outside the door and we will meet you.
Sample pick up is also available! Please email your name, location and number of samples to pbongior@bidmc.harvard.edu.
Pricing
Please call or email the facility for pricing information. Discounts are available for high volume projects!
Results
Each sample generates 2 files: the .seq text file and the .ab1 chromatogram.
The data can be obtained by emai OR posting on the DNASeqCore folder in the R drive (accessible only to the BIDMC community).
Accessing your results in the R drive
- You will need to map a network drive as follows:
- right click on “my computer”
- left click on “map network drive”
- choose drive letter “R”
- enter network address: \\sr42bromley\dnaseqcore
- Access the DNA Sequencing Core folder by logging in with the user id and password with which you log into your BIDMC computer.
- Select your PI’s folder.
- Select your folder.
- If you do not have access to the R drive, please contact me and I will arrange with IT to grant you access.
Viewing your results
The following are links to two sites from where you can download free software to view your samples’ .ab1 files:
Troubleshooting
Expert assistance is available in troubleshooting samples’ data and results optimization.
